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1.
Nat Commun ; 11(1): 5182, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33057025

RESUMO

Chronic obstructive pulmonary disease (COPD), diagnosed by reduced lung function, is a leading cause of morbidity and mortality. We performed whole genome sequence (WGS) analysis of lung function and COPD in a multi-ethnic sample of 11,497 participants from population- and family-based studies, and 8499 individuals from COPD-enriched studies in the NHLBI Trans-Omics for Precision Medicine (TOPMed) Program. We identify at genome-wide significance 10 known GWAS loci and 22 distinct, previously unreported loci, including two common variant signals from stratified analysis of African Americans. Four novel common variants within the regions of PIAS1, RGN (two variants) and FTO show evidence of replication in the UK Biobank (European ancestry n ~ 320,000), while colocalization analyses leveraging multi-omic data from GTEx and TOPMed identify potential molecular mechanisms underlying four of the 22 novel loci. Our study demonstrates the value of performing WGS analyses and multi-omic follow-up in cohorts of diverse ancestry.


Assuntos
Afro-Americanos/genética , Loci Gênicos , Doença Pulmonar Obstrutiva Crônica/genética , Fenômenos Fisiológicos Respiratórios/genética , Sequenciamento Completo do Genoma , Adulto , Idoso , Idoso de 80 Anos ou mais , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Proteínas de Ligação ao Cálcio/genética , Estudos de Viabilidade , Feminino , Seguimentos , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Pulmão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Proteínas Inibidoras de STAT Ativados/genética , Doença Pulmonar Obstrutiva Crônica/etnologia , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/genética
2.
Nat Commun ; 11(1): 5400, 2020 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-33106502

RESUMO

Slug (SNAI2), a member of the well-conserved Snail family of transcription factors, has multiple developmental roles, including in epithelial-to-mesenchymal transition (EMT). Here, we show that Slug is critical for the pathological angiogenesis needed to sustain tumor growth, and transiently necessary for normal developmental angiogenesis. We find that Slug upregulation in angiogenic endothelial cells (EC) regulates an EMT-like suite of target genes, and suppresses Dll4-Notch signaling thereby promoting VEGFR2 expression. Both EC-specific Slug re-expression and reduced Notch signaling, either by γ-secretase inhibition or loss of Dll4, rescue retinal angiogenesis in SlugKO mice. Conversely, inhibition of VEGF signaling prevents excessive angiogenic sprouting of Slug overexpressing EC. Finally, endothelial Slug (but not Snail) is activated by the pro-angiogenic factor SDF1α via its canonical receptor CXCR4 and the MAP kinase ERK5. Altogether, our data support a critical role for Slug in determining the angiogenic response during development and disease.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Células Endoteliais/metabolismo , Neovascularização Patológica/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Proteínas de Ligação ao Cálcio/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neovascularização Patológica/genética , Receptores Notch/genética , Receptores Notch/metabolismo , Transdução de Sinais , Fatores de Transcrição da Família Snail/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética
3.
BMC Med Genet ; 21(1): 177, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32887565

RESUMO

BACKGROUND: Keratoconus (KC) is usually bilateral, noninflammatory progressive corneal ectasia in which the cornea becomes progressively thin and conical, resulting in myopia, irregular astigmatism, and corneal scarring. METHODS: Eight families characterized by consanguineous marriages and/or multiple keratoconic individuals were examined genetically. Whole exome sequencing was done as trio or quadro per family. The output of the filtration procedure, based on minor allele frequency (MAF) less than 0.01 for homozygous variants and MAF equals 0 for heterozygous variants, is 22 missense variants. RESULTS: Based on the gene/protein function five candidate variants were highlighted in four families. Two variants were highlighted in one family within the genes MYOF and STX2, and one variant is highlighted in each of the other three families within the genes: COL6A5, ZNF676 and ZNF765. CONCLUSION: This study is one of the very rare that highlights genetic variants in association with KC.


Assuntos
Predisposição Genética para Doença/genética , Ceratocone/genética , Mutação , Sequenciamento Completo do Exoma/métodos , Proteínas de Ligação ao Cálcio/genética , Colágeno Tipo VI/genética , Consanguinidade , Proteínas de Ligação a DNA/genética , Saúde da Família , Feminino , Humanos , Jordânia , Ceratocone/diagnóstico , Masculino , Proteínas de Membrana/genética , Proteínas Musculares/genética , Linhagem , Sintaxina 1/genética
4.
Nat Commun ; 11(1): 3866, 2020 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-32737287

RESUMO

Upon severe head injury (HI), blood vessels of the meninges and brain parenchyma are inevitably damaged. While limited vascular regeneration of the injured brain has been studied extensively, our understanding of meningeal vascular regeneration following head injury is quite limited. Here, we identify key pathways governing meningeal vascular regeneration following HI. Rapid and complete vascular regeneration in the meninges is predominantly driven by VEGFR2 signaling. Substantial increase of VEGFR2 is observed in both human patients and mouse models of HI, and endothelial cell-specific deletion of Vegfr2 in the latter inhibits meningeal vascular regeneration. We further identify the facilitating, stabilizing and arresting roles of Tie2, PDGFRß and Dll4 signaling, respectively, in meningeal vascular regeneration. Prolonged inhibition of this angiogenic process following HI compromises immunological and stromal integrity of the injured meninges. These findings establish a molecular framework for meningeal vascular regeneration after HI, and may guide development of wound healing therapeutics.


Assuntos
Traumatismos Craniocerebrais/genética , Células Endoteliais/metabolismo , Neovascularização Fisiológica/genética , Regeneração/genética , Transdução de Sinais/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Vasos Sanguíneos/metabolismo , Vasos Sanguíneos/patologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Circulação Cerebrovascular , Traumatismos Craniocerebrais/metabolismo , Traumatismos Craniocerebrais/patologia , Modelos Animais de Doenças , Células Endoteliais/patologia , Regulação da Expressão Gênica/genética , Humanos , Macrófagos/metabolismo , Macrófagos/patologia , Meninges/lesões , Meninges/metabolismo , Camundongos , Camundongos Knockout , Receptor beta de Fator de Crescimento Derivado de Plaquetas/genética , Receptor beta de Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Cicatrização/genética
5.
Proc Natl Acad Sci U S A ; 117(36): 22204-22213, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32848060

RESUMO

The peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) is a transcriptional coactivator that controls expression of metabolic/energetic genes, programming cellular responses to nutrient and environmental adaptations such as fasting, cold, or exercise. Unlike other coactivators, PGC-1α contains protein domains involved in RNA regulation such as serine/arginine (SR) and RNA recognition motifs (RRMs). However, the RNA targets of PGC-1α and how they pertain to metabolism are unknown. To address this, we performed enhanced ultraviolet (UV) cross-linking and immunoprecipitation followed by sequencing (eCLIP-seq) in primary hepatocytes induced with glucagon. A large fraction of RNAs bound to PGC-1α were intronic sequences of genes involved in transcriptional, signaling, or metabolic function linked to glucagon and fasting responses, but were not the canonical direct transcriptional PGC-1α targets such as OXPHOS or gluconeogenic genes. Among the top-scoring RNA sequences bound to PGC-1α were Foxo1, Camk1δ, Per1, Klf15, Pln4, Cluh, Trpc5, Gfra1, and Slc25a25 PGC-1α depletion decreased a fraction of these glucagon-induced messenger RNA (mRNA) transcript levels. Importantly, knockdown of several of these genes affected glucagon-dependent glucose production, a PGC-1α-regulated metabolic pathway. These studies show that PGC-1α binds to intronic RNA sequences, some of them controlling transcript levels associated with glucagon action.


Assuntos
Glucagon/metabolismo , Glucagon/farmacologia , Hepatócitos/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Células Cultivadas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Gluconeogênese/fisiologia , Glucose/metabolismo , Guanosina Trifosfato/metabolismo , Fígado/metabolismo , Masculino , Metabolômica , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Transporte da Membrana Mitocondrial/genética , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Ligação Proteica , Transcriptoma
6.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 37(8): 828-832, 2020 Aug 10.
Artigo em Chinês | MEDLINE | ID: mdl-32761588

RESUMO

OBJECTIVE: To explore the genetic basis for a child with concomitant spinal muscular atrophy (SMA) and Citrin protein deficiency. METHODS: The child was subjected to whole exome sequencing by using target sequence capture high-throughput sequencing. Candidate variants were verified by Sanger sequencing. The SMN genes of the patient were also analyzed through multiplex ligation-dependent probe amplification (MLPA). RESULTS: The patient was found to carry homozygous deletion of exons 7 and 8 of the SMN1 gene, for which his parents were both carriers. The patient also carried compound heterozygous variants c.1737G>A and IVS16ins3kbof the SLA25A13 gene, in addition with compound heterozygous variants c.948G>A and c.2693T>C of the POLG gene, for which his parents were carriers, too. CONCLUSION: Variants of the SLC25A13 gene probably underlay the deficiency of Citrin protein, which may lead to neonatal intrahepatic cholestasis (NICCD). The patient also had SMA. The compound heterozygous variants c.948G>A and c.2693T>C of the POLG gene are likely to cause mitochondrial DNA deletion syndrome type 4A, though other types of mitochondrial disease cannot be excluded.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Atrofia Muscular Espinal/genética , Transportadores de Ânions Orgânicos/genética , Deficiência de Proteína/genética , Proteínas de Ligação ao Cálcio/deficiência , Criança , Polimerase do DNA Mitocondrial/genética , Homozigoto , Humanos , Proteínas de Transporte da Membrana Mitocondrial/genética , Transportadores de Ânions Orgânicos/deficiência , Deleção de Sequência , Proteína 1 de Sobrevivência do Neurônio Motor/genética
7.
Medicine (Baltimore) ; 99(27): e20995, 2020 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-32629717

RESUMO

RATIONAL: Intestinal lymphangiectasia (IL) is a rare disease characterized by dilatation and rupture of intestinal lymphatic channels leading to protein-losing enteropathy. IL is classified as primary and secondary types. PATIENT CONCERNS: A 3-month-old girl born at term from vaginal delivery with an APGAR score of 10/10 and birth weight of 4.310 g (>97° percentile) was admitted to our hospital because of increasing abdominal tenderness and diarrhea. At first examination, she presented an abdominal circumference of 60 cm, edema of the lower extremities and vulva, and facial dysmorphisms (hypertelorism, flat nasal bridge, flat mid-face). DIAGNOSIS: Once admitted, ultrasonography showed a large amount of ascites, while blood laboratory investigations revealed severe hypoproteinemia, hypoalbuminemia and hypogammaglobulinemia. Lymphoscintigraphy with 99m-Tc-nanocolloid demonstrated abnormal leakage of the tracer in the abdomen as evidence of IL. To detect a possible secondary, exams were performed and demonstrated positive antibody titres for CMV-IgM and IgG in blood and CMV-DNA positivity in blood, urine, saliva, maternal milk, and gastric and duodenal biopsies. Genetic investigations identified the genomic variant c.472C>T of the CCBE1 gene, coding for a protein variant (p.Arg158Cys), in homozygosity. INTERVENTIONS: Total parenteral nutrition was started and continued for a total of 18 days, then gradually bridged by enteral nutrition with a special formula. In addition, antiviral therapy for CMV infection was added first with intravenous ganciclovir for 14 days, resulting in the disappearance of blood viral load after 7 days of therapy and then with valganciclovir per os for another 30 days. OUTCOMES: The clinical course of the child gradually improved. A few days after starting treatments, lower extremities and vulvar edema disappeared, and abdominal circumference gradually decreased to a stable value of 38 cm, without any ultrasonographic signs of ascites left. Moreover, serum albumin and IgG rose to normal values after 3 months (4.3 g/dL and 501 mg/dL, respectively). LESSONS: This case suggests that in presence of IL both primary and secondary causes should be evaluated. On the other hand, genetic diagnosis is crucial not only for diagnosis but also for prognosis in HS. Life expectancy and quality could deeply vary among different gene mutations and protein variants of the same gene. Further studies and case reports are needed to better understand the clinical meaning of these genetic results and the role of CMV as trigger of IL.


Assuntos
Anormalidades Craniofaciais/genética , Linfangiectasia Intestinal/genética , Linfedema/genética , Proteínas de Ligação ao Cálcio/genética , Anormalidades Craniofaciais/diagnóstico , Feminino , Humanos , Lactente , Linfangiectasia Intestinal/diagnóstico , Linfedema/diagnóstico , Mutação , Proteínas Supressoras de Tumor/genética
8.
Rev Assoc Med Bras (1992) ; 66(5): 680-686, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32638975

RESUMO

OBJECTIVE Pelvic organ prolapse (POP) is a very frequent situation in our population that may lead to a significant decrease in patients' quality of life. Currently, we are looking for predictive factors for the development of POPs; thus, this study seeks to evaluate whether the Fibulin 5 polymorphism (FBLN5) is associated with the occurrence of POP. METHODS This is a cohort study with postmenopausal women who were divided into groups by POP stage: POP stages 0 and I (control group) and POP stages III and IV (case group). Subsequently, analyses of genetic polymorphisms of FBLN5 were performed using the Restriction Fragment Length Polymorphism (RFLP) technique. RESULTS A total of 292 women were included in the study. Pregnancy, parity and vaginal delivery in the patients, as well as in data described in the literature, were related to the occurrence of POP in the univariate analysis. However, after binary logistic regression, home birth and age remained independent risk factors for POP. We found no association between the FBLN5 polymorphism and the occurrence of POP (p = 0.371). CONCLUSION There was no association between the FBLN5 polymorphism and the occurrence of POP in Brazilian women.


Assuntos
Proteínas da Matriz Extracelular/genética , Prolapso de Órgão Pélvico , Qualidade de Vida , Brasil , Proteínas de Ligação ao Cálcio/genética , Estudos de Coortes , Feminino , Humanos , Polimorfismo Genético , Gravidez
9.
Tumour Biol ; 42(7): 1010428320937863, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32686600

RESUMO

Maintaining intracellular pH is crucial for preserving healthy cellular behavior and, when dysregulated, results in increased proliferation, migration, and invasion. The Na+/H+ exchanger isoform 1 is a highly regulated transmembrane antiporter that maintains pH homeostasis by exporting protons in response to intra- and extracellular signals. Activation of Na+/H+ exchanger isoform 1 is exquisitely regulated by the extracellular environment and protein cofactors, including calcineurin B homologous proteins 1 and 2. While Na+/H+ exchanger isoform 1 and calcineurin B homologous protein 1 are ubiquitously expressed, calcineurin B homologous protein 2 shows tissue-specific expression and upregulation in a variety of cancer cells. In addition, calcineurin B homologous protein 2 expression is modulated by tumorigenic extracellular conditions like low nutrients. To understand the role of calcineurin B homologous protein 2 in tumorigenesis and survival in lung cancer, we surveyed existing databases and formed a comprehensive report of Na+/H+ exchanger isoform 1, calcineurin B homologous protein 1, and calcineurin B homologous protein 2 expression in diseased and non-diseased tissues. We show that calcineurin B homologous protein 2 is upregulated during oncogenesis in many adeno and squamous carcinomas. To understand the functional role of calcineurin B homologous protein 2 upregulation, we evaluated the effect of Na+/H+ exchanger isoform 1 and calcineurin B homologous protein 2 depletion on cellular function during cancer progression in situ. Here, we show that calcineurin B homologous protein 2 functions through Na+/H+ exchanger isoform 1 to effect cell proliferation, cell migration, steady-state pHi, and anchorage-independent tumor growth. Finally, we present evidence that calcineurin B homologous protein 2 depletion in vivo has potential to reduce tumor burden in a xenograft model. Together, these data support the tumor-promoting potential of aberrant calcineurin B homologous protein 2 expression and position calcineurin B homologous protein 2 as a potential therapeutic target for the treatment of non-small cell lung cancer.


Assuntos
Calcineurina/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Trocador 1 de Sódio-Hidrogênio/metabolismo , Animais , Calcineurina/genética , Proteínas de Ligação ao Cálcio/genética , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Humanos , Concentração de Íons de Hidrogênio , Camundongos , Camundongos Nus , Transplante de Neoplasias , Isoformas de Proteínas/metabolismo , Transplante Heterólogo
10.
PLoS Biol ; 18(7): e3000782, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32692742

RESUMO

Tight regulation of gene transcription and mRNA splicing is essential for plant growth and development. Here we demonstrate that a plant-specific protein, EMBRYO DEFECTIVE 1579 (EMB1579), controls multiple growth and developmental processes in Arabidopsis. We demonstrate that EMB1579 forms liquid-like condensates both in vitro and in vivo, and the formation of normal-sized EMB1579 condensates is crucial for its cellular functions. We found that some chromosomal and RNA-related proteins interact with EMB1579 compartments, and loss of function of EMB1579 affects global gene transcription and mRNA splicing. Using floral transition as a physiological process, we demonstrate that EMB1579 is involved in FLOWERING LOCUS C (FLC)-mediated repression of flowering. Interestingly, we found that EMB1579 physically interacts with a homologue of Drosophila nucleosome remodeling factor 55-kDa (p55) called MULTIPLE SUPPRESSOR OF IRA 4 (MSI4), which has been implicated in repressing the expression of FLC by forming a complex with DNA Damage Binding Protein 1 (DDB1) and Cullin 4 (CUL4). This complex, named CUL4-DDB1MSI4, physically associates with a CURLY LEAF (CLF)-containing Polycomb Repressive Complex 2 (CLF-PRC2). We further demonstrate that EMB1579 interacts with CUL4 and DDB1, and EMB1579 condensates can recruit and condense MSI4 and DDB1. Furthermore, emb1579 phenocopies msi4 in terms of the level of H3K27 trimethylation on FLC. This allows us to propose that EMB1579 condensates recruit and condense CUL4-DDB1MSI4 complex, which facilitates the interaction of CUL4-DDB1MSI4 with CLF-PRC2 and promotes the role of CLF-PRC2 in establishing and/or maintaining the level of H3K27 trimethylation on FLC. Thus, we report a new mechanism for regulating plant gene transcription, mRNA splicing, and growth and development.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Proteínas de Ligação ao Cálcio/metabolismo , Desenvolvimento Vegetal/genética , Processamento de RNA/genética , Transcrição Genética , Proteínas de Arabidopsis/genética , Proteínas de Ligação ao Cálcio/genética , Núcleo Celular/metabolismo , Flores/fisiologia , Histonas/metabolismo , Mutação com Perda de Função , Lisina/metabolismo , Metilação , Proteínas Nucleares/metabolismo , Fenótipo , Raízes de Plantas/citologia , Ligação Proteica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sequências Repetitivas de Aminoácidos
11.
Arterioscler Thromb Vasc Biol ; 40(9): 2212-2226, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32640908

RESUMO

OBJECTIVE: The ductus arteriosus (DA) is a fetal artery connecting the aorta and pulmonary arteries. Progressive matrix remodeling, that is, intimal thickening (IT), occurs in the subendothelial region of DA to bring anatomic DA closure. IT is comprised of multiple ECMs (extracellular matrices) and migrated smooth muscle cells (SMCs). Because glycoprotein fibulin-1 binds to multiple ECMs and regulates morphogenesis during development, we investigated the role of fibulin-1 in DA closure. Approach and Results: Fibulin-1-deficient (Fbln1-/-) mice exhibited patent DA with hypoplastic IT. An unbiased transcriptome analysis revealed that EP4 (prostaglandin E receptor 4) stimulation markedly increased fibulin-1 in DA-SMCs via phospholipase C-NFκB (nuclear factor κB) signaling pathways. Fluorescence-activated cell sorting (FACS) analysis demonstrated that fibulin-1 binding protein versican was derived from DA-endothelial cells (ECs). We examined the effect of fibulin-1 on directional migration toward ECs in association with versican by using cocultured DA-SMCs and ECs. EP4 stimulation promoted directional DA-SMC migration toward ECs, which was attenuated by either silencing fibulin-1 or versican. Immunofluorescence demonstrated that fibulin-1 and versican V0/V1 were coexpressed at the IT of wild-type DA, whereas 30% of versican-deleted mice lacking a hyaluronan binding site displayed patent DA. Fibulin-1 expression was attenuated in the EP4-deficient mouse (Ptger4-/-) DA, which exhibits patent DA with hypoplastic IT, and fibulin-1 protein administration restored IT formation. In human DA, fibulin-1 and versican were abundantly expressed in SMCs and ECs, respectively. CONCLUSIONS: Fibulin-1 contributes to DA closure by forming an environment favoring directional SMC migration toward the subendothelial region, at least, in part, in combination with EC-derived versican and its binding partner hyaluronan.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Permeabilidade do Canal Arterial/metabolismo , Canal Arterial/metabolismo , Células Endoteliais/metabolismo , Matriz Extracelular/metabolismo , Miócitos de Músculo Liso/metabolismo , Animais , Proteínas de Ligação ao Cálcio/deficiência , Proteínas de Ligação ao Cálcio/genética , Movimento Celular , Células Cultivadas , Técnicas de Cocultura , Canal Arterial/anormalidades , Permeabilidade do Canal Arterial/genética , Permeabilidade do Canal Arterial/patologia , Células Endoteliais/patologia , Matriz Extracelular/genética , Matriz Extracelular/patologia , Humanos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos de Músculo Liso/patologia , NF-kappa B/metabolismo , Técnicas de Cultura de Órgãos , Proteína Quinase C/metabolismo , Ratos Wistar , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Transdução de Sinais , Fosfolipases Tipo C/metabolismo
12.
Yi Chuan ; 42(7): 703-712, 2020 Jul 20.
Artigo em Chinês | MEDLINE | ID: mdl-32694109

RESUMO

The analysis of genomic point mutations is one of the research strategies to explore the clonal evolution of tumor cells. At present, clonal evolution of tumor cells is mainly determined by bulk sampling and sequencing of different sections of the tumor. Since this approach analyzes a mixture of different cell types, it may not accurately represent the clonal evolution of specific tumor cell populations and likely miss low frequency mutations, especially when the sequencing depths are not sufficient. To address this issue, we have developed a strategy to analyze genomic point mutations from prostate basal cell carcinoma (BCC) tissues at single-cell resolution. Firstly, we optimized the single-cell whole genome amplification procedure with HepG2 cells. Then the single cells from BCC tissue were captured by a microfluidic chip of Fluidigm and processed for whole-genome amplification. Both SCUBE3 and MST1L genomic mutations were obtained by whole exome sequencing. Finally, we examined the genomic mutations through single-cell targeted amplification and Sanger sequencing. The established method successfully reconfirmed the mutations of SCUBE3 and MST1L in BCC at single cell level. The strategy established in this study could provide a useful tool for determining the clonal evolution of tumor cells based on genomic mutations at single-cell resolution.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Mutação Puntual , Projetos de Pesquisa , Proteínas de Ligação ao Cálcio/genética , Genoma , Genômica , Humanos , Masculino , Mutação , Receptores Proteína Tirosina Quinases/genética , Análise de Célula Única
13.
PLoS One ; 15(6): e0235122, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584873

RESUMO

The MGP single nucleotide polymorphism (SNP) rs1800801 has previously been associated with recurrent ischemic stroke in a Spanish cohort. Here, we tested for association of this SNP with ischemic stroke recurrence in a North American Caucasian cohort. Acute ischemic stroke patients admitted between 10/2009 and 12/2016 at three hospitals within a large healthcare system in the northeastern United States that were enrolled in a healthcare system-wide exome sequencing program were retrospectively reviewed. Patients with recurrent stroke within 1 year after index event were compared to those without recurrence. Of 9,348 suspected acute ischemic strokes admitted between 10/2009 and 12/2016, 1,727 (18.5%) enrolled in the exome-sequencing program. Among those, 1,068 patients had exome sequencing completed and were eligible for inclusion. Recurrent stroke within the first year of stroke was observed in 79 patients (7.4%). In multivariable analysis, stroke prior to the index stroke (OR 9.694, 95% CI 5.793-16.224, p ≤ 0.001), pro-coagulant status (OR = 3.563, 95% CI 1.504-8.443, p = 0.004) and the AA genotype of SNP rs1800801 (OR = 2.408, 95% CI 1.079-4.389, p = 0.004) were independently associated with recurrent stroke within the first year. The AA genotype of the MGP SNP rs1800801 is associated with recurrence within the first year after ischemic stroke in North American Caucasians. Study of stroke subtypes and additional populations will be required to determine if incorporation of allelic status at this SNP into current risk scores improves prediction of recurrent ischemic stroke.


Assuntos
Isquemia Encefálica/genética , Proteínas de Ligação ao Cálcio/genética , Proteínas da Matriz Extracelular/genética , Predisposição Genética para Doença , Genótipo , Polimorfismo de Nucleotídeo Único , Acidente Vascular Cerebral/genética , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva , Fatores de Risco , Sequenciamento Completo do Exoma
14.
Gen Physiol Biophys ; 39(3): 229-237, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32525816

RESUMO

For the exploration of circular RNA light chain kinase (circRNA-MYLK), siRNA#1 and siRNA#2 targeting circRNA-MYLK as well as microRNA(miR)-145-5p inhibitor were transfected. Viability was valued with the CCK-8. The protein expression was examined relying on Western blot. The expression of circRNA-MYLK or miR-145-5p was tested depending on qRT-PCR. The apoptotic/migration/invasion rate was separately measured by the Annexin v-FITC/PI with flow cytometer or chambers assays. CircRNA-MYLK was overexpressed in tumor tissue. Silencing circRNA-MYLK induced the inhibitions of viability, invasion and migration, as well as the blocks of MEK/ERK and NF-κB cascades, however, silencing circRNA-MYLK led to provoking of apoptosis. Besides, circRNA-MYLK silencing stimulated the over-production of miR-145-5p, whose silencing abolished the effects of siRNA#1 and siRNA#2 of circRNA-MYLK on those factors above. The circRNA-MYLK had oncogenic roles via targeting miR-145-5p in the Hep-2 cell line via stimulating MEK/ERK and NF-κB cascades.


Assuntos
Proteínas de Ligação ao Cálcio/genética , Carcinogênese , Neoplasias Laríngeas/patologia , MicroRNAs/genética , Quinase de Cadeia Leve de Miosina/genética , RNA Circular/genética , Apoptose , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Inativação Gênica , Humanos , Neoplasias Laríngeas/genética , Invasividade Neoplásica , Transdução de Sinais
15.
Int J Hematol ; 112(3): 361-368, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32535855

RESUMO

Genomic characterization of patients with myeloproliferative neoplasms (MPN) may lead to better diagnostic classification, prognostic assessment, and treatment decisions. These goals are particularly important in myelofibrosis (MF). We performed target Next Generation Sequencing for a panel of 255 genes and Chromosome Microarray Analysis (CMA) in 27 patients with MF. Patients were classified according to genomic findings and we compared the performance of a personalized prognostication system with IPSS, MIPSS70 and MIPSS70 + v2. Twenty-six patients presented mutations: 11.1% had single driver mutations in either JAK2, CALR or MPL; 85.2% had mutations in non-restricted genes (median: 2 per patient). CMA was abnormal in 91.7% of the 24 cases with available data. Copy-Number-Neutral Loss-of-Heterozygosity was the most common finding (66.7%). Del13q was the most frequent copy number variation, and we could define a 2.4 Mb minimally affected region encompassing RB1, SUCLA2 and CLLS2 loci. The largest genomic subgroup consisted of patients with mutations in genes involved with chromatin organization and splicing control (40.7%) and the personalized system showed better concordance and accuracy than the other prognostic systems. Comprehensive genomic characterization reveals the striking genetic complexity of MF and, when combined with clinical data, led, in our cohort, to better prognostication performance.


Assuntos
Variações do Número de Cópias de DNA , Genômica , Transtornos Mieloproliferativos/genética , Mielofibrose Primária/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Brasil , Proteínas de Ligação ao Cálcio/genética , Calreticulina/genética , Moléculas de Adesão Celular/genética , Estudos de Coortes , Feminino , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Janus Quinase 2/genética , Perda de Heterozigosidade/genética , Masculino , Análise em Microsséries/métodos , Pessoa de Meia-Idade , Mutação , Transtornos Mieloproliferativos/classificação , Mielofibrose Primária/classificação , Prognóstico
16.
PLoS One ; 15(5): e0233717, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32469983

RESUMO

Metastasis is known as a key step in cancer recurrence and could be stimulated by multiple factors. Calumenin (CALU) is one of these factors which has a direct impact on cancer metastasis and yet, its underlined mechanisms have not been completely elucidated. The current study was aimed to identify CALU co-expressed genes, their signaling pathways, and expression status within the human cancers. To this point, CALU associated genes were visualized using the Cytoscape plugin BisoGenet and annotated with the Enrichr web-based application. The list of CALU related diseases was retrieved using the DisGenNet, and cancer datasets were downloaded from The Cancer Genome Atlas (TCGA) and analyzed with the Cufflink software. ROC curve analysis was used to estimate the diagnostic accuracy of DEGs in each cancer, and the Kaplan-Meier survival analysis was performed to plot the overall survival of patients. The protein level of the signature biomarkers was measured in 40 biopsy specimens and matched adjacent normal tissues collected from CRC and lung cancer patients. Analysis of CALU co-expressed genes network in TCGA datasets indicated that the network is markedly altered in human colon (COAD) and lung (LUAD) cancers. Diagnostic accuracy estimation of differentially expressed genes showed that a gene panel consisted of CALU, AURKA, and MCM2 was able to successfully distinguish cancer tumors from healthy samples. Cancer cases with abnormal expression of the signature genes had a significantly lower survival rate than other patients. Additionally, comparison of CALU, AURKA, and MCM2 proteins between healthy samples, early and advanced tumors showed that the level of these proteins was increased through normal-carcinoma transition in both types of cancers. These data indicate that the interactions between CALU, AURKA, and MCM2 has a pivotal role in cancer development, and thereby needs to be explored in the future.


Assuntos
Aurora Quinase A , Proteínas de Ligação ao Cálcio , Neoplasias do Colo , Bases de Dados de Ácidos Nucleicos , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares , Componente 2 do Complexo de Manutenção de Minicromossomo , Aurora Quinase A/biossíntese , Aurora Quinase A/genética , Proteínas de Ligação ao Cálcio/biossíntese , Proteínas de Ligação ao Cálcio/genética , Neoplasias do Colo/diagnóstico , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/mortalidade , Intervalo Livre de Doença , Feminino , Humanos , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/mortalidade , Masculino , Componente 2 do Complexo de Manutenção de Minicromossomo/biossíntese , Componente 2 do Complexo de Manutenção de Minicromossomo/genética , Metástase Neoplásica , Taxa de Sobrevida
17.
PLoS One ; 15(5): e0233422, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32437418

RESUMO

SPARCL1 is a matricellular protein with anti-adhesive, anti-proliferative and anti-tumorigenic functions and is frequently downregulated in tumors such as colorectal carcinoma or non-small cell lung cancer. Studies have identified SPARCL1 as an angiocrine tumor suppressor secreted by tumor vessel endothelial cells, thereby exerting inhibitory activity on angiogenesis and tumor growth, in colorectal carcinoma. It is unknown whether SPARCL1 may exert these homeostatic functions in all organs and in other species. Therefore, SPARCL1 expression was comparatively analysed between humans and mice in a systematic manner. Murine Sparcl1 (mSparcl1) is most strongly expressed in the lung; expressed at an intermediate level in most organs, including the large intestine; and absent in the liver. In human tissues, SPARCL1 (hSPARCL1) was detected in all organs, with the strongest expression in the stomach, large intestine and lung, mostly consistent with the murine expression pattern. A striking difference between human and murine tissues was the absence of mSparcl1 expression in murine livers, while human livers showed moderate expression. Furthermore, mSparcl1 was predominantly associated with mural cells, whereas hSPARCL1 was detected in both mural and endothelial cells. Human SPARCL1 expression was downregulated in different carcinomas, including lung and colon cancers. In conclusion, this study revealed species-, organ- and cell-type-dependent expression of SPARCL1, suggesting that its function may not be similar between humans and mice.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Mucosa Intestinal/metabolismo , Fígado/metabolismo , Pulmão/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas da Matriz Extracelular/genética , Mucosa Gástrica , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Knockout , Especificidade de Órgãos , Especificidade da Espécie
18.
Nucleic Acids Res ; 48(12): 6839-6854, 2020 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-32449937

RESUMO

SERRATE/ARS2 is a conserved RNA effector protein involved in transcription, processing and export of different types of RNAs. In Arabidopsis, the best-studied function of SERRATE (SE) is to promote miRNA processing. Here, we report that SE interacts with the nuclear exosome targeting (NEXT) complex, comprising the RNA helicase HEN2, the RNA binding protein RBM7 and one of the two zinc-knuckle proteins ZCCHC8A/ZCCHC8B. The identification of common targets of SE and HEN2 by RNA-seq supports the idea that SE cooperates with NEXT for RNA surveillance by the nuclear exosome. Among the RNA targets accumulating in absence of SE or NEXT are miRNA precursors. Loss of NEXT components results in the accumulation of pri-miRNAs without affecting levels of miRNAs, indicating that NEXT is, unlike SE, not required for miRNA processing. As compared to se-2, se-2 hen2-2 double mutants showed increased accumulation of pri-miRNAs, but partially restored levels of mature miRNAs and attenuated developmental defects. We propose that the slow degradation of pri-miRNAs caused by loss of HEN2 compensates for the poor miRNA processing efficiency in se-2 mutants, and that SE regulates miRNA biogenesis through its double contribution in promoting miRNA processing but also pri-miRNA degradation through the recruitment of the NEXT complex.


Assuntos
Proteínas de Arabidopsis/genética , MicroRNAs/genética , RNA Helicases/genética , Processamento Pós-Transcricional do RNA/genética , Proteínas de Ligação a RNA/genética , Arabidopsis/genética , Proteínas de Ligação ao Cálcio/genética , Núcleo Celular/genética , Exossomos/genética , Regulação da Expressão Gênica de Plantas/genética , Mutação/genética , Precursores de RNA/genética , Estabilidade de RNA/genética , Ribonuclease III/genética
19.
PLoS Pathog ; 16(5): e1008214, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32379809

RESUMO

Calcium signaling plays a key role in many essential processes in almost all eukaryotic systems. It is believed that it may also be an important signaling system of the protist parasite Entamoeba histolytica. Motility, adhesion, cytolysis, and phagocytosis/trogocytosis are important steps in invasion and pathogenesis of E. histolytica, and Ca2+ signaling is thought to be associated with these processes leading to tissue invasion. There are a large number of Ca2+-binding proteins (CaBPs) in E. histolytica, and a number of these proteins appear to be associated with different steps in pathogenesis. The genome encodes 27 EF-hand-containing CaBPs in addition to a number of other Ca2+-binding domain/motif-containing proteins, which suggest intricate calcium signaling network in this parasite. Unlike other eukaryotes, a typical calmodulin-like protein has not been seen in E. histolytica. Though none of the CaBPs display sequence similarity with a typical calmodulin, extensive structural similarity has been seen in spite of lack of significant functional overlap with that of typical calmodulins. One of the unique features observed in E. histolytica is the identification of CaBPs (EhCaBP1, EhCaBP3) that have the ability to directly bind actin and modulate actin dynamics. Direct interaction of CaBPs with actin has not been seen in any other system. Pseudopod formation and phagocytosis are some of the processes that require actin dynamics, and some of the amoebic CaBPs (EhC2Pk, EhCaBP1, EhCaBP3, EhCaBP5) participate in this process. None of these E. histolytica CaBPs have any homolog in organisms other than different species of Entamoeba, suggesting a novel Ca2+ signaling pathway that has evolved in this genus.


Assuntos
Cálcio/metabolismo , Entamoeba histolytica/metabolismo , Entamebíase/metabolismo , Actinas/metabolismo , Cálcio/fisiologia , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Calmodulina/metabolismo , Entamoeba histolytica/genética , Entamoeba histolytica/patogenicidade , Fagocitose , Proteínas de Protozoários/metabolismo
20.
Am J Ophthalmol ; 218: 40-53, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32445700

RESUMO

PURPOSE: To characterize the progression of optical gaps and expand the known etiologies of this phenotype. DESIGN: Retrospective cohort study. METHODS: Thirty-six patients were selected based on the identification of an optical gap on spectral-domain optical coherence tomography (OCT) from a large cohort of patients (N = 746) with confirmed diagnoses of inherited retinal dystrophy. The width and height of the gaps in 70 eyes of 36 patients were measured by 2 independent graders using the caliper tool on Heidelberg Explorer. Measurements of outer and central retinal thickness were also evaluated and correlated with gap dimensions. RESULTS: Longitudinal analysis confirmed the progressive nature of optical gaps in patients with Stargardt disease, achromatopsia, occult macular dystrophy, and cone dystrophies (P < .003). Larger changes in gap width were noted in patients with Stargardt disease (78.1 µm/year) and cone dystrophies (31.9 µm/year) compared with patients with achromatopsia (16.2 µm/year) and occult macular dystrophy (15.4 µm/year). Gap height decreased in patients with Stargardt disease (6.5 µm/year; P = .02) but increased in patients with achromatopsia (3.3 µm/year) and occult macular dystrophy (1.2 µm/year). Gap height correlated with measurements of central retinal thickness at the fovea (r = 0.782, P = .00012). Interocular discordance of the gap was observed in 7 patients. Finally, a review of all currently described etiologies of optical gap was summarized. CONCLUSION: The optical gap is a progressive phenotype seen in an increasing number of etiologies. This progressive nature suggests a use as a biomarker in the understanding of disease progression. Interocular discordance of the phenotype may be a feature of Stargardt disease and cone dystrophies.


Assuntos
Biomarcadores , Defeitos da Visão Cromática/diagnóstico por imagem , Distrofias de Cones e Bastonetes/diagnóstico por imagem , Degeneração Macular/diagnóstico por imagem , Retinite Pigmentosa/diagnóstico por imagem , Doença de Stargardt/diagnóstico por imagem , Tomografia de Coerência Óptica , Adolescente , Adulto , Idoso , Proteínas de Ligação ao Cálcio/genética , Criança , Defeitos da Visão Cromática/fisiopatologia , Distrofias de Cones e Bastonetes/fisiopatologia , Progressão da Doença , Eletrorretinografia , Feminino , Humanos , Degeneração Macular/fisiopatologia , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Fenótipo , Retina/fisiopatologia , Retinite Pigmentosa/genética , Retinite Pigmentosa/fisiopatologia , Estudos Retrospectivos , Doença de Stargardt/fisiopatologia , Acuidade Visual/fisiologia , Proteínas rab de Ligação ao GTP/genética
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